Heterologous UTR sequences for enhanced mRNA expression

Abstract

mRNAs containing an exogenous open reading frame (ORF) flanked by a 5′ untranslated region (UTR) and a 3′ UTR is provided, wherein the 5′ and 3′ UTRs are derived from a naturally abundant mRNA in a tissue. Also provided are methods for identifying the 5′ and 3′ UTRs, and methods for making and using the mRNAs.

What is claimed is: 
     
       1. A method of expressing a protein in a liver tissue, comprising providing an mRNA to said liver tissue, wherein the mRNA comprises an open reading frame (ORF) that encodes said protein, flanked by a heterologous 5′ untranslated region (UTR) and/or a heterologous 3′ UTR, wherein the 5′ UTR and/or the 3′ UTR are derived from a gene selected from the group consisting of Haptoglobin (HP), Haptoglobin-Related Protein (HPR), Complement Component 3 (C3), Alpha-1-antitrypsin (SERPINA1), Alpha-1-antichymotrypsin (SERPINA3), Fibrinogen Alpha Chain (FGA), and Fibrinogen Beta Chain (FGB). 
     
     
       2. The method of  claim 1 , wherein the mRNA comprises at least one modified or non-naturally occurring nucleotide. 
     
     
       3. The method of  claim 2 , wherein the at least one modified or non-naturally occurring nucleotide comprises at least one backbone modification, sugar modification or base modification. 
     
     
       4. The method of  claim 3 , wherein the at least one modified or non-naturally occurring nucleotide comprises at least one base modification. 
     
     
       5. The method of  claim 4 , wherein the at least one base modification is selected from the group consisting of: 2-amino-6-chloropurine riboside 5′ triphosphate, 2-aminoadenosine 5′ triphosphate, 2-thiocytidine 5′ triphosphate, 2-thiouridine 5′ triphosphate, 4-thiouridine 5′ triphosphate, 5-aminoallylcytidine 5′ triphosphate, 5-aminoallyluridine 5′ triphosphate, 5-bromocytidine 5′ triphosphate, 5-bromouridine 5′ triphosphate, 5-iodocytidine 5′ triphosphate, 5-iodouridine 5′ triphosphate, 5-methylcytidine 5′ triphosphate, 5-methyluridine 5′ triphosphate, 6-azacytidine 5′ triphosphate, 6-azauridine 5′ triphosphate, 6-chloropurine riboside 5′-triphosphate, 7-deazaadenosine 5′ triphosphate, 7-deazaguanosine 5′ triphosphate, 8-azaadenosine 5′ triphosphate, 8-azidoadenosine 5′ triphosphate, benzimidazole riboside 5′ triphosphate, N 1 -methyladenosine 5′ triphosphate, N 1 -methylguanosine 5′ triphosphate, N 6 -methyladenosine 5′ triphosphate, O 6 -methylguanosine 5′ triphosphate, N 1 -methyl-pseudouridine 5′ triphosphate, puromycin 5′-triphosphate and xanthosine 5′ triphosphate. 
     
     
       6. The method of  claim 5 , wherein the at least one modified or non-naturally occurring nucleotide is N 1 -methyl-pseudouridine 5-triphosphate. 
     
     
       7. The method of  claim 1 , wherein the mRNA comprises a heterologous 5′ UTR derived from a gene selected from the group consisting of: HP, FGB, HPR, C3, FGA, SERPINA1 and SERPINA3. 
     
     
       8. The method of  claim 1 , wherein the 5′ UTR comprises a nucleotide sequence selected from the group consisting of: SEQ ID NOS:1-3, 5-7, and 10. 
     
     
       9. The method of  claim 1 , wherein the ORF is therapeutic gene. 
     
     
       10. The method of  claim 9 , wherein the therapeutic gene is selected from the group consisting of: interferon alpha, TNF related apoptosis-inducing ligand, vascular adhesion protein 1, hepatocyte growth factor, G6PC, ABCB11, ABCB4, ASL1, ASS, Arg1, AGXT, OTC, CBS, PAH, GBE, GALE, HADH, MCCC1, MMA, ADAMTS13, SLC25A15, C2, F2, PROS1, SERPINA1, GALT1, ETFA, GCDH, CTNS, FAH, TAT, HPD, HSD17b4, SLC2A2, GALC, ABCC6, AHSG, PDC-E2, GAA, ATP8B1, MMACHC, GK, PCCA, PCCB and UGT1A1. 
     
     
       11. The method of  claim 1 , wherein the mRNA comprises a heterologous 5′ UTR derived from the FGB gene. 
     
     
       12. The method of  claim 1 , wherein the 5′ UTR comprises the nucleotide sequence set forth in SEQ ID NO:2.